Main Cardiac Histopathologic Alterations in the Acute Phase of Trypanosoma cruzi Infection in a Murine Model

Abstract

Symptoms in the acute phase of Chagas disease are usually mild and nonspecific. However, after several years, severe complications like dilated heart failure and even death may arise in the chronic phase. Due to the lack of specific symptoms in the acute phase, the aim of this work was to describe and analyze the cardiac histopathology during this phase in a CD1 mouse model by assessing parasitism, fibrotic damage, and the presence and composition of a cellular infiltrate, to determine its involvement in the pathogenesis of lesions in the cardiac tissue. Our results indicate that the acute phase lasts about 62 days post-infection (dpi). A significant increase in parasitemia was observed since 15 dpi, reaching a maximum at 33 dpi (4.1 × 10⁶). The presence of amastigote nests was observed at 15-62 dpi, with a maximum count of 27 nests at 35 dpi. An infiltrate consisting primarily of macrophages and neutrophils was found in the cardiac tissue within the first 30 days, but the abundance of lymphocytes showed an 8 ≥ fold increase at 40-62 dpi. Unifocal interstitial fibrosis was identified after 9 dpi, which subsequently showed a 16 ≥ fold increase at 40-60 dpi, along with a 50% mortality rate in the model under study. The increased area of fibrotic lesions revealed progression in the extent of fibrosis, mainly at 50-62 dpi. The presence of perivasculitis and thrombus circulation disorders was seen in the last days (62 dpi); finally, cases of myocytolysis were observed at 50 and 62 dpi. These histopathological alterations, combined with collagen deposition, seem to lead to the development of interstitial fibrosis and damage to the cardiac tissue during the acute phase of infection. This study provides a more complete understanding of the patterns of histopathological abnormalities involved in the acute phase, which could help the development of new therapies to aid the preclinical tests of drugs for their application in Chagas disease.

Keywords: acute chagas disease; cardiac histopathology; mouse model; pathogenicity.

Figure 1

(A). Parasitemia, and (B). Parasitism. (A). Number of parasites. 95% CI of median. Confidence level = 98.08%, 5 replicates of dpi. (B). In amastigote nests, CI of median. Two types of nests were found in the myocardium: (a) surrounded by infiltrate, and (b) without inflammatory infiltrate. 95% CI of median. Actual confidence level, 98.08%. 5 replicates. Inoculum: 1000 parasites. N = 80 mice.

Figure 2

Photomicrographs of cardiac tissue at different times post-infection during experimental murine acute Chagas disease (4000×). H.E.—H.E. stain. M—Masson’s stain. 0 dpi (A,B)—myocardium-septum: no amastigote nests; normal cardiac histology, no infiltrate, and no lesions. 9 dpi, myocardium-apex: (C)—mixed infiltrate (arrow); (D)—unifocal lesion with scant infiltrate (arrow). 15 dpi, myocardium-septum: (E)—mixed infiltrate (arrow); (F)—irregular interstitial fibrosis with scant infiltrate (arrow). 19 dpi, myocardium-right ventricle: (G)—mixed interstitial and lymphocytic infiltrate in myocardium and endocardium (arrow); (H)—interstitial fibrosis with adjacent infiltrate (arrow). 27 dpi, myocardium-left ventricle: (I)—intense inflammatory infiltrate (arrow); (J)—multifocal interstitial fibrosis (arrow).

Figure 3

Photomicrographs of heart at 33 dpi. Image at 1000× (left) H.E.—H.E. stain; M—Masson’s stain. (A,B)—decreased apex is shown, and several areas with interstitial fibrosis (rectangle); (4000× right, rectangle zoom-in) (C)— mixed infiltrate, H.E.; (D)—presence of extensive interstitial fibrosis.

Figure 4

Photomicrographs of myocardium at different times post-infection (4000×) H.E.—H.E. stain. M—Masson’s stain. 36 dpi, myocardium-left ventricle: (A)—amastigote nest (arrow). (B)—amastigote nest (arrow). 39 dpi, right myocardium-ventricle: (C)—amastigote nest and lymphocytic infiltrate (arrows) (1000×). (D)—amastigote nest (arrow) (E)—43 dpi, intracavitary thrombus proximal to the apex (arrow); 43 dpi (4000×) (F,G)—perivasculitis, fibrosis, and infiltrate (arrow).

Figure 5

Photomicrographs of mouse myocardium at different times post-infection (4000×). H.E.—H.E. stain. M—Masson’s stain. 57 dpi, myocardium-septum: (A)— lymphocyte infiltrate (arrow); (B)—lymphocyte infiltrate and interstitial fibrosis (arrow); (C,D)— vacuolization and myocytolysis (arrows); (E,F)—62 dpi, endocardium between septum and left ventricle (1000×): intracavitary thrombi in both ventricles (arrow). 62 dpi (4000×), (G,H)—septum, lymphocytic infiltrate (arrows).

Figure 6

Prevalence of infiltrate in the experimental acute phase of Chagas disease. Mixed-lymphocyte infiltrate with few macrophages and neutrophils. Lymphocytic-only lymphocytes. The increase in lymphocytic infiltrate with respect to mixed infiltrate is noteworthy. * Presence of perivasculitis.

Figure 7

Prevalence and location of infiltrate in heart structures in the experimental acute phase of Chagas disease. M, myocardium; M + EP, myocardium and epicardium; M + EN, myocardium and endocardium; P(M + EN + EP) myocardium, epicardium, and endocardium or pancarditis.

Figure 8

Prevalence of interstitial fibrosis in the experimental acute phase of Chagas disease. FI-UF, unifocal interstitial fibrosis; FI-MF, multifocal interstitial fibrosis; FI-E, extensive interstitial fibrosis. Mortality at 60 dpi; 16-fold increase in FI.5, with 50% mortality in the study model.

Figure 9

Increase of the areas of fibrotic interstitial lesions at 0–60 dpi in the acute phase. 9 dpi, apex, unifocal lesion. 20 dpi, left ventricle, multifocal lesion. 40 dpi, septum, extensive lesion. 50 dpi, septum, multifocal lesions. 60 dpi, septum, lesions. Differences were analyzed by bilateral unpaired t-test. p = 0.0033.

Figure 10

Model for the presence of histopathological alterations in acute murine experimental T. cruzi infection. Summary of the progression of histopathological lesions observed. dpi, days post-infection.